Bottom Line:
The pathogenesis of F. tularensis is not fully understood, and research has failed to develop an effective vaccine to this date.Furthermore, we demonstrate that targeting inactivated F. tularensis (iFt) to Fcγ receptors (FcɣRs) via intranasal immunization with mAb-iFt complexes, a proven vaccine strategy in our laboratories, reverses the anti-inflammatory effects of the bacterium on macrophages by down-regulating production of IL-10.These results provide important insight for further understanding the protective immune mechanisms generated when targeting immunogens to Fc receptors.

Affiliation: Department of Biological Sciences, Seton Hall University, South Orange, New Jersey, United States of America.

ABSTRACTProduction of pro-inflammatory cytokines by innate immune cells at the early stages of bacterial infection is important for host protection against the pathogen. Many intracellular bacteria, including Francisella tularensis, the agent of tularemia, utilize the anti-inflammatory cytokine IL-10, to evade the host immune response. It is well established that IL-10 has the ability to inhibit robust antigen presentation by dendritic cells and macrophages, thus suppressing the generation of protective immunity. The pathogenesis of F. tularensis is not fully understood, and research has failed to develop an effective vaccine to this date. In the current study, we hypothesized that F. tularensis polarizes antigen presenting cells during the early stages of infection towards an anti-inflammatory status characterized by increased synthesis of IL-10 and decreased production of IL-12p70 and TNF-α in an IFN-ɣ-dependent fashion. In addition, F. tularensis drives an alternative activation of alveolar macrophages within the first 48 hours post-infection, thus allowing the bacterium to avoid protective immunity. Furthermore, we demonstrate that targeting inactivated F. tularensis (iFt) to Fcγ receptors (FcɣRs) via intranasal immunization with mAb-iFt complexes, a proven vaccine strategy in our laboratories, reverses the anti-inflammatory effects of the bacterium on macrophages by down-regulating production of IL-10. More specifically, we observed that targeting of iFt to FcγRs enhances the classical activation of macrophages not only within the respiratory mucosa, but also systemically, at the early stages of infection. These results provide important insight for further understanding the protective immune mechanisms generated when targeting immunogens to Fc receptors.

pone.0129981.g002: Immunization with mAb-iFt immune complexes increases the activation of PECs following LVS challenge.C57BL/6 mice were immunized i.n. with PBS, iFt (2x107 CFUs), or mAb-iFt, boosted on day 21 and challenged on day 35 with 10,000 CFUs of Ft LVS. On day 2 post-infection the peritoneal exudate cells of immunized mice were harvested and the expression of F4/80, MHC class II, B7.1 (CD80), and B7.2 (CD86) were analyzed by flow cytometry. Results are representative of three independent experiments. (*) P-value < 0.1; (**) P-value < 0.05; bars represent the SD.

Mentions:
Peritoneal cells were obtained from immunized mice two days post-LVS challenge as described in Materials and Methods, and the expression of the murine macrophage cell-surface marker F4/80, the co-stimulatory molecules CD80 and CD86, as well as MHC class II was determined by flow cytometry. Although the number of cells expressing the F4/80 cell surface marker was similar between the iFt and mAb-iFt immunized mice, the number and frequency of cells expressing both MHC class II and CD80/CD86 molecules was significantly increased in the mAb-iFt group (Fig 2). This enhancement in surface marker expression upon immunization with mAb-iFt correlates with the increased presentation of iFt in the presence of mAb-iFt in vitro [26].

pone.0129981.g002: Immunization with mAb-iFt immune complexes increases the activation of PECs following LVS challenge.C57BL/6 mice were immunized i.n. with PBS, iFt (2x107 CFUs), or mAb-iFt, boosted on day 21 and challenged on day 35 with 10,000 CFUs of Ft LVS. On day 2 post-infection the peritoneal exudate cells of immunized mice were harvested and the expression of F4/80, MHC class II, B7.1 (CD80), and B7.2 (CD86) were analyzed by flow cytometry. Results are representative of three independent experiments. (*) P-value < 0.1; (**) P-value < 0.05; bars represent the SD.

Mentions:
Peritoneal cells were obtained from immunized mice two days post-LVS challenge as described in Materials and Methods, and the expression of the murine macrophage cell-surface marker F4/80, the co-stimulatory molecules CD80 and CD86, as well as MHC class II was determined by flow cytometry. Although the number of cells expressing the F4/80 cell surface marker was similar between the iFt and mAb-iFt immunized mice, the number and frequency of cells expressing both MHC class II and CD80/CD86 molecules was significantly increased in the mAb-iFt group (Fig 2). This enhancement in surface marker expression upon immunization with mAb-iFt correlates with the increased presentation of iFt in the presence of mAb-iFt in vitro [26].

Bottom Line:
The pathogenesis of F. tularensis is not fully understood, and research has failed to develop an effective vaccine to this date.Furthermore, we demonstrate that targeting inactivated F. tularensis (iFt) to Fcγ receptors (FcɣRs) via intranasal immunization with mAb-iFt complexes, a proven vaccine strategy in our laboratories, reverses the anti-inflammatory effects of the bacterium on macrophages by down-regulating production of IL-10.These results provide important insight for further understanding the protective immune mechanisms generated when targeting immunogens to Fc receptors.

Affiliation:
Department of Biological Sciences, Seton Hall University, South Orange, New Jersey, United States of America.

ABSTRACTProduction of pro-inflammatory cytokines by innate immune cells at the early stages of bacterial infection is important for host protection against the pathogen. Many intracellular bacteria, including Francisella tularensis, the agent of tularemia, utilize the anti-inflammatory cytokine IL-10, to evade the host immune response. It is well established that IL-10 has the ability to inhibit robust antigen presentation by dendritic cells and macrophages, thus suppressing the generation of protective immunity. The pathogenesis of F. tularensis is not fully understood, and research has failed to develop an effective vaccine to this date. In the current study, we hypothesized that F. tularensis polarizes antigen presenting cells during the early stages of infection towards an anti-inflammatory status characterized by increased synthesis of IL-10 and decreased production of IL-12p70 and TNF-α in an IFN-ɣ-dependent fashion. In addition, F. tularensis drives an alternative activation of alveolar macrophages within the first 48 hours post-infection, thus allowing the bacterium to avoid protective immunity. Furthermore, we demonstrate that targeting inactivated F. tularensis (iFt) to Fcγ receptors (FcɣRs) via intranasal immunization with mAb-iFt complexes, a proven vaccine strategy in our laboratories, reverses the anti-inflammatory effects of the bacterium on macrophages by down-regulating production of IL-10. More specifically, we observed that targeting of iFt to FcγRs enhances the classical activation of macrophages not only within the respiratory mucosa, but also systemically, at the early stages of infection. These results provide important insight for further understanding the protective immune mechanisms generated when targeting immunogens to Fc receptors.